A cold-adapted (ca) reassortant influenza B virus vaccine was restricted in replication in the upper and lower respiratory tract of hamsters and chimpanzees compared to its wild type influenza B virus parent. Isolates of the influenza B ca reassortant recovered after up to 15 days of replication in immunosuppressed hamsters did not exhibit instability of the ts or attenuation phenotype as indicated by tests performed in cell culture and in hamsters. Attempts are underway to isolate phenotypic revertants of previously characterized influenza A virus reassortants which derived only one gene from the influenza A/Ann Arbor/60 ca donor virus, while the remaining genes were derived from a wild type influenza A virus. Each of four ts+ revertants of a single gene PB2 reassortant virus also regained the ability to replicate in the lower respiratory tract of hamsters indicating that attenuation specified by the ca PB2 gene is closely linked to the ts phenotype. Phenotypic revertants of single gene PB1 and M reassortant viruses have also been isolated and are currently under evaluation. The NS gene of avian influenza A viruses exhibits sequence dimorphism, i.e., there are two allelic forms of the NS gen, allele A or allele B. One of these alleles (A) is also present in human influenza A viruses, while the other avian influenza A virus Ns allele (B) is highly divergent from the corresponding gene of the human influenza A viruses. The level of replication of these reassortant viruses in squirrel monkeys will be compared with that of a human wild type influenza A virus to determine if the B allele specifies host range restriction in primate respiratory tissue. Phenotypic and genotypic stability are important properties of the M and NP genes of the avian influenza A/Mallard/78 donor virus that play a role in host-range restriction of avian-human influenza A virus vaccines in primates. Both phenotypic and genotypic stability of these genes are being studied by biological and sequence analysis of single M or NP gene reassortants which have been passaged serially in monkeys.